New players in the regulation of ecdysone biosynthesis

Insect ecdysone steroid hormone regulates major developmental transitions, such as molting and metamorphosis. The production of ecdysone correlates well with the timing of these transitions. Finding out how the ecdysone biosynthesis is regulated is crucial to fully understand these sophisticated developmental switches. Here we summarized recent findings in the regulation of ecdysone biosynthesis from the aspects of cell signaling, key biosynthetic enzymes and substrate cholesterol trafficking.

Effects of β-ecdysone and polypodine-B on moulting and growth of prawn, Penaeus orientalis

Effects of the mixture of β-ecdysone and polypodine -B (2 : 1 W/W) on the moulting and growth of prawn, Penaeus orientaliss were studied in laboratory by rearing the animal with the added compound diets. The results showed that the addition of the moulting hormone (M. H) mixture to compound diets, from 1. 33× 10-6 to 30×10-6, could increase moulting activity and growth rate of the prawn to different extents, among which the 5. 33× 10-6-10. 67×10-6addition had statistically significant effects. However, 60 × 10-6 added diets caused moulting and growth inhibitions and cuticle thickening. The analyses of protein, water and ash contents of the prawn showed no remarkable differences between hormone dosed animals and the control.

Genome-wide CRISPR screening reveals key genes and pathways associated with 20-hydroxyecdysone signal transduction in the silkworm(Bombyx mori)

Metamorphosis is a complex developmental process involving multiple pathways and a large number of genes that are regulated by juvenile hormone(JH)and 20-hydroxyecdysone(20E).Despite important progress in understanding various aspects of silkworm biology,the hormone signaling pathway in the silkworm remains poorly understood.Genome-wide screening using clustered regularly interspaced short palindromic repeats(CRISPR)/CRISPR-associated protein 9(Cas9)-based libraries has recently emerged as a novel method for analyzing genome function,enabling further research into essential genes,drug targets,and virus-host interaction.Previously,we constructed a genome-wide CRISPR/Cas9-based library of the silkworm(Bombyx mori)and successfully revealed the genes involved in biotic or abiotic stress factor responses.In this study,we used our silkworm CRISPR library and large-scale genome-wide screening to analyze the key genes in the silkworm 20E signaling pathway and their mechanisms of action.Functional annotation showed that 20E regulates key proteins in processes that mainly occur in the cytoplasm and nucleus.Pathway enrichment analysis showed that 20E can activate phosphorylation and may affect innate immunity,interfere with intracellular nutrition and energy metabolism,and eventually cause cell apoptosis.The screening results were experimentally validated by generating cells with knockout alleles of the relevant genes,which had increased tolerance to 20E.Our findings provide a panoramic overview of signaling in response to 20E in the silkworm,underscoring the utility of genome-wide CRISPR mutant libraries in deciphering hormone signaling pathways and the mechanisms that regulate metamorphosis in insects.

Simultaneous determination of six steroidal saponins and one ecdysone in Asparagus filicinus using high performance liquid chromatography coupled with evaporative light scattering detection

A high-performance liquid chromatography coupled with an evaporative light scattering detector(HPLC-ELSD)has been developed to evaluate the quality of Asparagus filicinus through a simultaneous determination of six steroidal saponins and one ecdysone,including aspafiliosides A,B,C,E,G,filiasparoside A and 20-hydroxyecdysone.With a C_(18) analytical column,the seven analytes were separated efficiently using acetonitrile-water as the mobile phase in a gradient program.The method limits of detection ranged 0.1250.225μg,and the method limits of quantitation ranged 0.408-0.720μg,respectively.The intra-and inter-day precisions of the method were evaluated and were all less than 3%.All the recoveries for the spiked analytes ranged 95.16%-100.61%.The proposed method was succesfully applied to quantify the seven components in thirteen samples from different localities in China.

MicroRNA targets the ecdysone signaling pathway E75 gene to control larval-pupal development in Bactrocera dorsalis

MicroRNAs (miRNAs) regulate various biological processes during insect developme nt;however, their role in larval-pupal development in oriental fruit fly, Bactrocera dorsalis (Hendel) remains unknown. In the current study, we address the biological function of a conserved miRNA, Bdo-Let-7 in the regulation of BdE75 gene, which belongs to the ecdysone signaling pathway and participates in the larval-pupal development in B. dorsalis. Using dual luciferase reporter assay in HEK293T cells we show that Bdo-Let-7 miRNA interacts with the 3' untranslated region of BdE75 gene and suppresses its expression. The Bdo-Let?7 and BdE75 are also co-expressed in the larval-pupal stages and in different tissues of B. dorsalis .In in vivo experiments, the injectio n of Bdo-Let-7 agomir and antagomir in third instar larvae down- and up-regulated the expression of BdE75、 respectively. The 20-hydroxyecdysone (20E) injection assay shows that 20E up-regulated the expression of Bdo-Let-7 on the 5th day of the larvae. Moreover, abnormal pupation and eclosion were observed after larval Bdo-Let-7 antagomir injection. Based on these results, we show that Bdo-Let-7 regulates the ecdysone signaling pathway through the exact dose of BdE75 gene, and is indispensable for normal larval-pupal development in B. dorsalis.

Multiple microRNAs control ecdysone signaling in the midgut of Spodoptera litura

Metamorphosis is one of the most important physiological processes in insects.It is regulated by a serial of ecdysone cascade genes.Recently,lots o f microRNAs(miRNAs)were investigated in insects;however,their function in metamorphosis is largely unknown.In the present study,the dynamics of a small RNA population was investigated by RNA sequencing from the midgut of a lepidopteran pest Spodoptera litura during larval-pupal metamorphosis.A total of 101 miRNAs were identified,and 75 miRNAs were differentially expressed during the metamorphic process.The relationship between these differentially expressed miRNAs and 12 ecdysone cascade genes was analyzed by four classical software programs,and a multiple-to-multiple regulatory network was found to exist between these miRNAs and their targets.Among them,miR-14-3p and its two targets(EcR and E75)were chosen for further validation.MiR-14-3p had higher expression level in the 6 th instar larvae as compared with either the prepupae or pupae,which was opposite to that of both EcR and E75,two ecdysone cascade genes.Luciferase reporter assay confirmed that both EcR and E75 were regulated by miR-14-3p.Interestingly,the 3'untranslated regions are nearly identical to each other among different transcript variants of the ecdysone cascade genes,including EcR,USP,E75,E74,E78,E93,Hr3,Hr4,Hr39,K rhl and F tzfl.Thus,different transcript variants of one ecdysone cascade gene could be regulated by the same mi RNA.The above data suggest that the ecdysone signaling pathway is under the tight control of miRNA.These findings expand our understanding of the mechanism of insect metamorphosis and may also provide a novel possibility for the control of pest insects in the future.

RNA interference targeting ecdysone receptor blocks the larval-pupal transition in Henosepilachna vigintioctopunctata

Henosepilachna vigintioctopunctata is a serious insect pest which attacks a large number of nightshades and cucurbits in Asian countries,Brazil and Australia.Prolonged application of traditional pesticides has caused environmental pollution and exerted deleterious effects on human health.Finding new approaches with high target specificity and low environmental contamination has become an urgent task.RNA interference(RNAi)induced by double-stranded RNA(dsRNA)is expected to be applicable to managing this pest.Here we evaluated the effects of Escherichia co/Z-expressed dsRNAs targeting ecdvsone receptor(EcR)gene via dietary delivery in laboratory and foliar spraying in a greenhouse.The target transcript was successfully knocked down when the 4th-instar larvae had fed on potatofoliage dipped with dsEcR in a laboratory bioassay.Around 85%of the HvEcR RNAi larvae remained as prepupae or became abnormal pupae,and failed to emerge into adults.Ingestion of ds£c7?-immersed foliage by the 3rd-instar larvae effectuated a comparable RNAi response and brought about more severe defects:all the resultant larvae arrested development,remained as prepupae and finally died.For assay in the greenhouse,a ds£c7?-contained E.coli suspension was directly sprayed to the foliage of greenhouse-growing potato plants and the 3rd-and 4th-instar larvae were transferred to the leaves.High RNAi efficacy was obtained and identical RNAi phenotypes were observed in treated larvae.In addition,spraying dsEcR reduced leaf damage.Our results indicate a possibility of practical application of dsEcR as an environmentally friendly RNA pesticide to control H.vigintioctopunctata larvae.

Ecdysone receptor isoforms play distinct roles in larval–pupal–adult transition in Leptinotarsa decemlineata

A heterodimer of two nuclear receptors,ecdysone receptor(EcR)and ultraspiracle,mediates 20‐hydroxyecdysone(20E)signaling to modulate many aspects in insect life,such as molting and metamorphosis,reproduction,diapause and innate immunity.In the present paper,we intended to determine the isoform‐specific roles of EcR during larval–pupal–adult transition in the Colorado potato beetle.Double‐stranded RNAs(dsRNAs)were prepared using the common(dsEcR)or isoform‐specific(dsEcRA,dsEcRB1)regions of EcR as templates.Ingestion of either dsEcR or dsEcRA,rather than dsEcRB1,by the penultimate(3rd)and final(4th)instar larvae caused failure of larval–pupal and pupal–adult ecdysis.The RNA interference(RNAi)larvae remained as prepupae,or became deformed pupae and adults.Determination of messenger RNA(mRNA)levels of EcR isoforms found that LdEcRA regulates the expression of LdEcRB1.Moreover,silencing the two EcR transcripts,LdEcRA or LdEcRB1 reduced the mRNA levels of Ldspo and Ldsad,and lowered 20E titer.In contrast,the expression levels of HR3,HR4,E74 and E75 were significantly decreased in the LdEcR or LdEcRA RNAi larvae,but not in LdEcRB1 depleted specimens.Dietary supplement with 20E did not restore the expression of five 20E signaling genes(USP,HR3,HR4,E74 and E75),and only partially alleviated the pupation defects in dsEcR‐or dsEcRA‐fed beetles.These data suggest that EcR plays isoform‐specific roles in the regulation of ecdysteroidogenesis and the transduction of 20E signal in L.decemlineata.

Ecdysone and Insulin Signaling Play Essential Roles in Readjusting the Altered Body Size Caused by the dGPAT4 Mutation in Drosophila

Body size is one of the features that distinguish one species from another in the biological world. Animals have developed mechanisms to control their body size during normal development. However, how animals cope with genetic alterations and/or environmental stresses to develop into normal-sized adults remain poorly understood. The ability of the animals to develop into a normal-sized adult after the challenges of genetic alterations and/or environmental stresses reveals a robustness of body size control. Here we show that the mutation of dGPAT4, a de novo synthase of lysophosphatidic acid, is a genetic alteration that triggers such a robust response of the animals to body size challenges in Drosophila. Loss of dGPAT4 leads to a severe delay of development, slow growth and resultant small-sized animals during the larval stages, but results in normal-sized adult flies. The robust body size adjustment of the dGPAT4 mutant is likely achieved by corresponding changes in ecdysone and insulin signaling, which is also manifested by compromised food intake. Thus, we propose that a strategy has been evolved by the animals to reach final body size when challenged by genetic alterations, which requires the coordinated ecdysone and insulin signaling.

Dietary cholesterol promotes growth and ecdysone signalling pathway by modulating cholesterol transport in swimming crabs(Portunus trituberculatus)

Cholesterol,as an indispensable nutrient,regulates molting and growth in crustacean.As crustaceans are unable to biosynthesize cholesterol de novo,it is central to understand how dietary cholesterol affects molting in crustaceans.An 8-week feeding trial was conducted to evaluate the effects of dietary cholesterol level(0.12%,0.43%,0.79%,1.00%,1.30%and 2.50%)on growth,cholesterol metabolism and expression of genes related to lipid and ecdysone metabolism in female swimming crabs(Portunus trituberculatus).A total of 192 crabs(1.41±0.05 g)were randomly distributed into 192 aquaria.Each treatment had 4 replicates with each replicate containing 8 crabs.Crabs fed the 1.00%cholesterol diet showed best growth performance,and thus based on percent weight gain,the optimal dietary cholesterol requirement was calculated at 1.01%.Tissue cholesterol concentrations were positively correlated with dietary cholesterol level.The contents of functional fatty acids in hepatopancreas significantly increased as dietary cholesterol increased from 0.12%to 2.50%(P<0.05).The expression levels of genes related to lipogenesis pathway,lipid catabolism and fatty acid oxidation were significantly downregulated with increased dietary cholesterol level(P<0.05).The highest expression levels of cholesterol transport genes,low-density lipoprotein receptor(ldlr)and low-density lipoprotein receptorrelated protein 2(lrp2)occurred in crabs fed the 1.30%cholesterol diet.Moreover,hormones related to molting such as crustacean hyperglycemic hormone(CHH),methyl farnesoate(MF),molt-inhibiting hormone(MIH),and ecdysone in hemolymph were significantly influenced by dietary cholesterol level(P<0.05).The highest expression levels of ecdysone receptor(ecr)and chitinase 1(chi1)in eyestalk and hepatopancreas were found in crabs fed the diet containing 1.00%cholesterol(P<0.05).In conclusion,the optimal dietary level was beneficial to functional fatty acid accumulation,regulated lipid metabolism,promoted the ecdysone signalling pathway by improving the cholesterol transport,and improved the molting rate and growth of swimming crabs.

The Ecdysone and Notch Pathways Synergistically Regulate Cut at the Dorsal-Ventral Boundary in Drosophila Wing Discs

Metazoan development requires coordination of signaling pathways to regulate patterns of gene expression.In Drosophila,the wing imaginal disc provides an excellent model for the study of how signaling pathways interact to regulate pattern formation.The determination of the dorsal-ventral（DV） boundary of the wing disc depends on the Notch pathway,which is activated along the DV boundary and induces the expression of the homeobox transcription factor,Cut.Here,we show that Broad（Br）,a zinc-finger transcription factor,is also involved in regulating Cut expression in the DV boundary region.However,Br expression is not regulated by Notch signaling in wing discs,while ecdysone signaling is the upstream signal that induces Br for Cut upregulation.Also,we find that the ecdysone-Br cascade upregulates cut-lacZ expression,a reporter containing a 2.7 kb cut enhancer region,implying that ecdysone signaling,similar to Notch,regulates cut at the transcriptional level.Collectively,our findings reveal that the Notch and ecdysone signaling pathways synergistically regulate Cut expression for proper DV boundary formation in the wing disc.Additionally,we show br promotes Delta,a Notch ligand,near the DV boundary to suppress aberrant high Notch activity,indicating further interaction between the two pathways for DV patterning of the wing disc.

荻草谷网蚜ABCG1基因的克隆及表达模式分析

荻草谷网蚜Sitobion miscanthi是严重威胁我国小麦生产安全的迁飞性害虫。蜕皮激素是参与蚜虫翅型分化调控的内激素,在有翅成蚜体内保持高滴度,且诱导后代产生更高比例的无翅蚜,其进出靶细胞需要经过细胞膜上特定蛋白的转运。ATP结合盒转运蛋白家族G亚家族(ATP-binding cassette transporter G, ABCG)中的ABCG1是通过跨膜转运昆虫类固醇、对蜕皮激素信号进行负调控的功能蛋白之一,在蚜虫中尚未见报道。本文克隆了荻草谷网蚜ABCG1(SmisABCG1)基因,并进行了序列比对、系统进化分析以及不同组织部位和发育时期表达模式分析。结果显示,SmisABCG1基因的开放阅读框全长为1 851 bp,编码616个氨基酸,含7个跨膜结构域,符合ABCG蛋白家族典型结构特性,基因登录ID:OP626323。昆虫间ABCG1较保守,该蛋白系统进化关系与各自物种间亲缘关系的远近保持一致。其中,SmisABCG1与来自豌豆蚜、禾谷缢管蚜、棉蚜、花生蚜和雪松长足大蚜等的ABCG1氨基酸序列高度一致(>87%),以上蚜虫聚为一支。与SmisABCG1亲缘关系最近的是豌豆蚜的ABCG1,其次是半翅目的褐飞虱、白背飞虱和灰飞虱,与膜翅目的新疆菜叶蜂、阿里山潜蝇茧蜂以及鞘翅目的赤拟谷盗、蜂箱小甲虫亲缘关系较远。该基因在伪胚胎和成蚜阶段高表达。包含伪胚胎的有翅、无翅成蚜整蚜SmisABCG1的转录水平无显著差异,但其在来自有翅成蚜的伪胚胎中的转录水平高于无翅成蚜伪胚胎,证实无翅成蚜自身的转录水平较高,而有翅成蚜较低。进一步分析显示这一差异主要是无翅蚜胸部显著高表达所导致。基于该蛋白对蜕皮激素负调控,与有翅成蚜转录水平低,但蜕皮激素水平更高相符合。

芒果点斑螟不同发育阶段转录组及保幼激素和蜕皮激素相关基因分析

【目的】探明芒果点斑螟(Citripestis eutraphera)不同发育阶段的转录组差异,挖掘保幼激素(JH)和蜕皮激素(Ecd)合成等生长发育相关基因,为芒果点斑螟特异性防治和研发环保型生长调节类杀虫剂打下基础。【方法】利用高通量测序技术对芒果点斑螟幼虫、蛹和成虫进行转录组测序、数据组装、功能注释和比较分析,筛选出JH和Ecd合成相关基因,并通过实时荧光定量PCR进行验证。【结果】芒果点斑螟幼虫、蛹和成虫3个阶段共组装得到254154条Unigenes。幼虫与蛹阶段比较筛选出10688个差异表达基因(DEGs),幼虫与成虫阶段比较筛选出10041个DEGs,蛹与成虫阶段比较筛选出12896个DEGs;将DEGs进行Nr数据库比对,结果显示与脐橙螟蛾(Amyelois transitella)的序列相似度最高;进行GO功能注释分析,共注释到61266条Unigenes,各发育阶段芒果点斑螟的DEGs多富集在细胞结构体、细胞过程、催化活性和结合等过程;在KEGG数据库中共有14731条Unigenes得到注释,涉及147个代谢通路,DEGs富集在代谢过程、核糖体和氧化磷酸化过程等通路的数目较多,筛选出49条Unigenes涉及JH和Ecd合成相关基因,对DIB、JHAMT、FPPP、ALDH-1和ALDH-2等5个DEGs进行实时荧光定量PCR分析,结果ALDH-1、ALDH-2DIB和DIB在蛹期高表达,且相对表达量显著高于其他时期(P<0.05,下同),FPPP和JHAMT在幼虫期低表达且相对表达量显著低于其他时期。【结论】基于转录组分析获得不同发育阶段芒果点斑螟的DEGs多富集在细胞结构体、细胞过程、催化活性和结合等过程,且主要富集在代谢相关通路中;筛选出49条与JH和Ecd合成相关的基因,DIB、JHAMT、FPPP、ALDH-1和ALDH-2等基因在芒果点斑螟不同发育阶段表达差异显著,可能在芒果点斑螟生长发育中发挥重要作用。

三七总皂苷复方脂质体凝胶剂的制备及皮肤给药研究

目的制备三七总皂苷复方脂质体凝胶剂,并进行体外透皮吸收研究。方法以薄膜分散法制备脂质体,通过正交实验设计优化三七总皂苷复方脂质体的处方,再以卡波姆为基质制成凝胶剂。通过智能透皮试验仪,进行体外透皮实验,研究三七总皂苷中人参皂苷Rg_1及β-蜕皮激素的体外透皮吸收情况,并采用HPLC法检测透过及滞留在皮肤内的Rg_1和β-蜕皮激素量。结果卵磷脂与胆固醇用量比为9:1,Rg_1质量浓度为40 g·L^(-1)β-蜕皮激素质量浓度为3g·L^(-1),脂药比为5:1时的处方为最佳脂质体处方,卡波姆-934重量比为1%时,药物的皮肤滞留量最大。结论将三七总皂苷制备成复方脂质体凝胶剂,能够增加药物的皮肤滞留量,提高药物在皮肤局部的生物利用度。

果蝇蜕皮激素诱导程序性细胞死亡的遗传调控因子

近年来关于果蝇程序性细胞死亡(programmedcelldeath,PCD)的研究结果表明,在果蝇的变态发育过程中,蜕皮激素与受体结合后诱导转录因子的表达。这些转录因子作为程序性细胞死亡调控网络中的初、次级应答信号,激活凋亡诱导因子Reaper、Hid和Grim的表达。Reaper、Hid和Grim进而阻止凋亡蛋白抑制因子的活性,从而启动半胱氨酸蛋白酶caspase途径,引起细胞凋亡(apoptosis)。该文综述了蜕皮激素诱导的果蝇程序性细胞死亡中各遗传调控因子之间的关系。

昆虫脑神经肽的研究进展:抑前胸腺肽PTSP

1999年发现和鉴定的新型昆虫脑神经肽——抑前胸腺肽 ,对昆虫的变态起着重要的作用 .本文介绍它的研究背景、结构、功能、与促前胸腺素的相互作用关系及其分子生物学研究的结果 。

江西青牛胆的化学成分

从江西青牛胆(TinosporacravenianaS .Y .Hu)块根中分到7个化合物,经波谱分析,这些化合物的化学结构分别确定为:古伦宾(columbin ,1)、异古伦宾(isocolumbin ,2 )、巴马亭(palmatine ,3)、β谷甾醇( βsitosterol,4 )、2 0 β羟基蜕皮激素( 2 0 βhydroxy ecdysone ,5 )、巴马土宾(palmatrubine ,6 )、2 0 β羟基蜕皮激素 2 O β葡萄糖甙( 2 0 βecdys terone 2 O βD glucopyranoside ,7)。

露水草β-蜕皮激素含量积累规律与最佳采收期研究

为确认露水草的最佳采收期,观测了露水草植株的生长发育进程,根及地上部分干物质和β-蜕皮激素的积累规律,结果表明:露水草出苗至抽薹期为缓慢生长阶段,开花期为其快速生长阶段,生育期195 d时干物质积累达到顶峰,结实后为稳定生长阶段;根的β-蜕皮激素含量是地上部分的4～20倍;生育期135 d后,根β-蜕皮激素含量逐渐增加,195 d时最高,以后快速下降。从而认为根为露水草的采收部位,生育期195 d是露水草的最佳采收期。

海南青牛胆非生物碱成分的分离与鉴定(Ⅱ)

从防己科植物的一个新种——海南青牛胆（ＴｉｎｏｓｐｏｒａＨａｉｎａｎｅｓｉｓＨ．Ｓ．ＬｏｅｔＺ．Ｘ．Ｌｉ）的藤茎中又分得２个非生物碱化合物，经光谱解析，鉴定为丁香甙（ｓｙｒｉｎｇｉｎ）和２０－羟基蜕皮素（２０－ｈｙ－ｄｒｏｘｙｅｃｄｙｓｏｎｅ）。

褐飞虱蜕皮激素合成相关Halloween基因的克隆及功能分析

【目的】蜕皮激素(ecdysone)是调控昆虫生长发育和繁殖的重要激素,其合成主要由5个称为Halloween基因编码的细胞色素P450(spook/CYP307A1,Phantom/CYP306A1,disembodied/CYP302A1,shadow/CYP315A1和shade/CYP314A1)参与。褐飞虱Nilaparvata lugens系东南亚和东亚地区危害最为严重的水稻害虫之一,以蜕皮激素调控基因为靶标,应用RNAi技术防控褐飞虱正逐步成为研究热点。本研究旨在揭示褐飞虱体内Halloween基因的功能,为褐飞虱的防控和新农药的开发提供理论依据。【方法】本研究基于褐飞虱基因组和转录组数据,利用RT-PCR技术克隆了参与蜕皮激素合成的5个Halloween基因;利用生物信息学软件预测分析其编码蛋白的结构特性;利用MEGA5.0邻接法构建系统进化树;利用实时荧光定量PCR技术研究其时空表达特征;利用注射法RNAi技术分析Cyp314a1基因表达下调对褐飞虱生长发育和繁殖的影响。【结果】克隆获得褐飞虱的5个Halloween基因,即NlCyp307a1(Gen Bank登录号:KM217014.1),NlCyp306a1(Gen Bank登录号:KM217013.1),NlCyp302a1(Gen Bank登录号:KM216995.1),NlCyp315a1(Gen Bank登录号:KM216998.1)和NlCyp314a1(Gen Bank登录号:KU928172)。进化分析表明,这5个基因分属于CYP2和线粒体两大类群,并与各自的同源基因聚类在一起,说明Halloween基因和蜕皮激素合成途径在昆虫进化过程中十分保守。Nl CYP314A1具有5个P450的保守结构域和两个跨膜结构域,是典型的线粒体酶。qRT-PCR检测结果显示,5个Halloween基因的表达量在整个褐飞虱5龄若虫期内呈现波动状,且均在5龄若虫蜕皮后的24 h和60 h出现表达量峰值。NlCyp314a1在褐飞虱发育各个龄期均表达,且在成虫中表达量最高;NlCyp314a1在胸部表达量较头部和腹部高,其在脂肪体中的表达量最高,其次是在足、表皮、翅芽中,中肠中表达量最低。利用注射法RNAi对NlCyp314a1基因进行沉默后,NlCyp314a1的表达量在注射后第4天较对照极显著降低84.6%(P<0.01),同时蜕皮激素通路下游的响应关键基因Nl FTZ-F1的表达量较对照极显著降低64.1%(P<0.01)。表型观察发现,NlCyp314a1表达量的下调导致了褐飞虱蜕皮困难,出现畸形虫体,第7天死亡率大于95%,且NlCyp314a1表达量的下调导致了褐飞虱卵巢发育畸形。【结论】褐飞虱蜕皮激素合成相关的Halloween基因在进化上非常保守,Halloween基因功能的研究揭示了褐飞虱体内Halloween基因对于褐飞虱蜕皮和繁殖的重要性。其表达水平的降低导致了褐飞虱生长发育和繁殖受阻,提示褐飞虱Halloween基因可用于褐飞虱蜕皮和繁殖进程的调控,同样也为Halloween基因在褐飞虱防治方面的潜在应用提供了依据。